Figure 4

Detection of the aph VIII gene in transformants. (A1-A3) Paromomycin-resistant transformants and the parental wild-type strain were analyzed for presence of the aphVIII gene in the genome by genomic PCR. The expected size of the aphVIII fragment produced in transformants was 324 bp (Figure 3A). The rightmost lane (pPmr3) shows a positive control using pPmr3 plasmid DNA as the template. Lane M refers to the molecular weight marker. (A4) Sequence obtained from the amplified and cloned aphVIII fragments. The positions of the primers and the start codon (bold) are indicated. (B) Southern blot analysis of HindIII- or BamHI-digested genomic DNA of two randomly chosen transformants, EuHR-8 and EuHR-12, and from the parental wild-type strain. The blot was probed using a 282-bp fragment of the aphVIII coding region (Figure 3A).